High frequency RNA recombination in porcine reproductive and respiratory syndrome virus occurs preferentially between parental sequences with high similarity

Two types of porcine reproductive and respiratory syndrome virus (PRRSV) exist, a North American type and a European type. The co-existence of both types in some countries, such as Denmark, Slovakia and Canada, creates a risk of inter-type recombination. To evaluate this risk, cell cultures were co-infected with either a North American and a European type of PRRSV or two diverse types of European isolate. Subsequently, an approximately 600 bp region of the PRRSV genome was tested for recombination by quantitative real-time RT–PCR. Between 0·1 and 2·5% RNA recombination was found between the European isolates, but no recombination was detected between the European and North American types. Calculation of the maximum theoretical risk of European–American recombination, based on the sensitivity of the RT–PCR system, revealed that RNA recombination between the European and North American types of PRRSV is at least 10000 times less likely to occur than RNA recombination between diverse European isolates.

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Sequencing and analysis of complete genome of the attenuated Hanvet1.VN strain used for vaccine production against porcine reproductive and respiratory syndrome

Vietnam Journal of Biotechnology ◽

10.15625/1811-4989/16/1/13168 ◽

2018 ◽

Vol 16 (1) ◽

pp. 51-57

Author(s):

Nguyễn Thị Nga ◽

Hà Thị Thu ◽

Nguyễn Thị Hoa ◽

Vũ Thị Hiền ◽

Trần Thị Thu Hiền ◽

...

Keyword(s):

North American ◽

Sanger Sequencing ◽

Respiratory Syndrome Virus ◽

Whole Genome ◽

Type Ii ◽

Vaccine Production ◽

Rt Pcr ◽

North American Strain ◽

American Strain ◽

Vietnamese Strain

The porcine reproductive and respiratory syndrome virus (PRRSV) attenuated strain Hanvet1.VN has been developed by the Pharmaceutical and Veterinary Material J.S.C (HANVET) by passaging HY-2010 strain on MARC-145 cells for 80 passages and used for PRRS vaccine production. In this study, we sequenced and analyzed the whole genome of the attenuated Hanvet1.VN strain. The total RNA was extracted from the Hanvet1.VN strain, RT-PCR was used for amplification of 15 separate segments of the whole genome. The amplified segments were cloned into the pCR2.1 vector and sequenced by Sanger sequencing. The sequences were analyzed with BioEdit and DNA Star Software. The results showed that, GP5 of the Hanvet1.VN attenuated strain had 100% identity in amino acid (aa) sequences with one of the pathogenic Vietnamese strain isolated in Quang Nam Province and had 98% identity with that of the Chinese 07NM strain. However, the identity of aa sequence of the Hanvet1.VN GP5 was much lower in the comparison with GP5 of VR2332, and it was only 87%. The MP and NP proteins were highly conserved compared with pathogenic strains circulating in Vietnam (07QN) and China (07NM) (99-100%, respectively). The other eight proteins of the Hanvet1.VN strain showed changes from 1.2% in NP1a to 3.9% in GP2 compared with the 07QN strain. However, the aa identity of all Hanvet1.VN proteins were very low when compared with proteins of PRRSV type II strain (North American strain, VR2332), ranged from 86.25% to 97.7%. Our results showed that the Hanvet1.VN attenuated vaccine strain had protective immunogenicity similar to that strain circulating in Vietnam closely related to a strain from China but different from the type II North American strain VR2332. Hence, for importing PRRSV vaccine, especially from American or Europe Countries, antigenic compatibility of the PRRSV vaccine and strains circulating in Vietnam should be concerned in PRRSV vaccine production.

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Phylogeny-Based Evolutionary, Demographical, and Geographical Dissection of North American Type 2 Porcine Reproductive and Respiratory Syndrome Viruses

Journal of Virology ◽

10.1128/jvi.02551-09 ◽

2010 ◽

Vol 84 (17) ◽

pp. 8700-8711 ◽

Cited By ~ 178

Author(s):

Mang Shi ◽

Tommy Tsan-Yuk Lam ◽

Chung-Chau Hon ◽

Michael P. Murtaugh ◽

Peter R. Davies ◽

...

Keyword(s):

United States ◽

North American ◽

Phylogenetic Analyses ◽

The United States ◽

Respiratory Syndrome Virus ◽

Viral Spread ◽

Comprehensive Picture ◽

North American Type ◽

Orf5 Sequences

ABSTRACT Type 2 (or North American-like) porcine reproductive and respiratory syndrome virus (PRRSV) was first recorded in 1987 in the United States and now occurs in most commercial swine industries throughout the world. In this study, we investigated the epidemiological and evolutionary behaviors of type 2 PRRSV. Based on phylogenetic analyses of 8,624 ORF5 sequences, we described a comprehensive picture of the diversity of type 2 PRRSVs and systematically classified all available sequences into lineages and sublineages, including a number of previously undescribed lineages. With the rapid growth of sequence deposition into the databases, it would be technically difficult for veterinary researchers to genotype their sequences by reanalyzing all sequences in the databases. To this end, a set of reference sequences was established based on our classification system, which represents the principal diversity of all available sequences and can readily be used for further genotyping studies. In addition, we further investigated the demographic histories of these lineages and sublineages by using Bayesian coalescence analyses, providing evolutionary insights into several important epidemiological events of type 2 PRRSV. Moreover, by using a phylogeographic approach, we were able to estimate the transmission frequencies between the pig-producing states in the United States and identified several states as the major sources of viral spread, i.e., “transmission centers.” In summary, this study represents the most extensive phylogenetic analyses of type 2 PRRSV to date, providing a basis for future genotyping studies and dissecting the epidemiology of type 2 PRRSV from phylogenetic perspectives.

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Simultaneous Detection of North American and European Porcine Reproductive and Respiratory Syndrome Virus Using Real-Time Quantitative Reverse Transcriptase–PCR

Journal of Veterinary Diagnostic Investigation ◽

10.1177/104063870501700211 ◽

2005 ◽

Vol 17 (2) ◽

pp. 165-170 ◽

Cited By ~ 57

Author(s):

Steven B. Kleiboeker ◽

Susan K. Schommer ◽

Sang-Myeong Lee ◽

Sandy Watkins ◽

Wayne Chittick ◽

...

Keyword(s):

Reverse Transcriptase ◽

Real Time ◽

North American ◽

Simultaneous Detection ◽

Viral Rna ◽

Respiratory Syndrome Virus ◽

Analytical Sensitivity ◽

Rt Pcr ◽

Field Isolates ◽

Pcr Assays

Porcine reproductive and respiratory syndrome (PRRS) is 1 of the most economically important diseases of swine. Detection of the etiologic agent, PRRS virus (PRRSV), represents a diagnostic challenge due to the heterogeneity of field isolates as well as the propensity for swine to develop persistent infection in which virus is difficult to detect. Recently European (EU) lineage PRRSV isolates, which are genetically divergent from North American (NA) isolates, have been introduced into NA swine further complicating efforts to diagnose this disease. In this study, real-time ( TaqMan) reverse transcriptase (RT)–PCR assays were developed for multiplex detection, differentiation, and quantification of NA and EU PRRSV field isolates. Oligonucleotide primers and dual-labeled probes were selected from conserved regions of open-reading frame 7 and the 3'-untranslated region. The real-time RT-PCR assays described for the NA or EU genotype of PRRSV detected viral RNA from 83/83 strains (74 NA; 9 EU) previously isolated by cell culture between 1992 and 2003. The analytical sensitivity of both assays was consistently found to be less than a single TCID50, which corresponded to 5–10 RNA molecules, and was not significantly reduced when the reactions were performed in a multiplex format. When performing multiplex reactions, sensitive detection was possible even when 1 viral RNA concentration was up to 5,000-fold higher than the second. The diagnostic sensitivity and specificity of the multiplex reaction was found to be at a minimum equivalent to that of both nested RT-PCR and virus isolation.

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Quantitative TaqMan® RT-PCR for the detection and differentiation of European and North American strains of porcine reproductive and respiratory syndrome virus

Journal of Virological Methods ◽

10.1016/s0166-0934(01)00358-5 ◽

2001 ◽

Vol 98 (1) ◽

pp. 63-75 ◽

Cited By ~ 52

Author(s):

Christoph Egli ◽

Barbara Thür ◽

Luzia Liu ◽

Martin A Hofmann

Keyword(s):

North American ◽

Respiratory Syndrome Virus ◽

Rt Pcr

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Genetic polymorphism of the nsp2 gene in North American type-Porcine reproductive and respiratory syndrome virus

Archives of Virology ◽

10.1007/s00705-008-0098-6 ◽

2008 ◽

Vol 153 (7) ◽

pp. 1323-1334 ◽

Cited By ~ 45

Author(s):

Masaaki Yoshii ◽

Tatsuyuki Okinaga ◽

Ayako Miyazaki ◽

Kanako Kato ◽

Hidetoshi Ikeda ◽

...

Keyword(s):

Genetic Polymorphism ◽

North American ◽

Respiratory Syndrome Virus ◽

Nsp2 Gene ◽

North American Type

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Simultaneous detection of Classical swine fever virus and North American genotype Porcine reproductive and respiratory syndrome virus using a duplex real-time RT-PCR

Journal of Virological Methods ◽

10.1016/j.jviromet.2008.05.011 ◽

2008 ◽

Vol 151 (2) ◽

pp. 194-199 ◽

Cited By ~ 16

Author(s):

Dan Cheng ◽

Jian-Jun Zhao ◽

Na Li ◽

Yuan Sun ◽

Yan-Jun Zhou ◽

...

Keyword(s):

Real Time ◽

North American ◽

Classical Swine Fever Virus ◽

Simultaneous Detection ◽

Classical Swine Fever ◽

Fever Virus ◽

Respiratory Syndrome Virus ◽

Rt Pcr ◽

North American Genotype

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A Full-Length cDNA Infectious Clone of North American Type 1 Porcine Reproductive and Respiratory Syndrome Virus: Expression of Green Fluorescent Protein in the Nsp2 Region

Journal of Virology ◽

10.1128/jvi.01032-06 ◽

2006 ◽

Vol 80 (23) ◽

pp. 11447-11455 ◽

Cited By ~ 96

Author(s):

Ying Fang ◽

Raymond R. R. Rowland ◽

Michael Roof ◽

Joan K. Lunney ◽

Jane Christopher-Hennings ◽

...

Keyword(s):

United States ◽

Green Fluorescent Protein ◽

North American ◽

Fluorescent Protein ◽

Infectious Clone ◽

The United States ◽

Respiratory Syndrome Virus ◽

Green Fluorescent ◽

North American Type

ABSTRACT The recent emergence of a unique group of North American type 1 porcine reproductive and respiratory syndrome virus (PRRSV) in the United States presents new disease control problems for a swine industry that has already been impacted seriously by North American type 2 PRRSV. In this study, a full-length cDNA infectious clone was generated from a low-virulence North American type 1 PRRSV isolate, SD01-08. In vitro studies demonstrated that the cloned virus maintained growth properties similar to those of the parental virus. Virological, pathological, and immunological observations from animals challenged with cloned viruses were similar to those from animals challenged with the parental virus and a modified live virus vaccine. To further explore the potential use as a viral backbone for expressing foreign genes, the green fluorescent protein (GFP) was inserted into a unique deletion site located at amino acid positions 348 and 349 of the predicted Nsp2 region in the virus, and expression of the Nsp2-GFP fusion protein was visualized by fluorescent microscopy. The availability of this North American type 1 infectious clone provides an important research tool for further study of the basic viral biology and pathogenic mechanisms of this group of type 1 PRRSV in the United States.

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